The major goal of this research is to delineate mechanisms whereby various nutritional factors and pharmacological agents influence the activation and conjugation of precarcinogens in intact cells, using isolated perfused ogans as experimental models. The following studies are proposed: (1) Investigate the influence of NADPH supply in rates of mixed-function oxidation in the isolated hemoglobin-free perfused rat liver and rabbit lung using benzo(a)pyrene 7-ethoxycoumarin and p-nitroanisol as model substrates. Alterations in rates of mixed-function oxidation that occur in the presence of specific inhibitors, inducing agents and alterations in nutritional status will be explored to determine the relative contribution of mitochondrial and cytosolic sources of NADPH and NADH. Steady-state concentrations of key intermediates of energy metabolism, pyridine nucleotides, oxidation-reduction induced changes in surface fluorescence of intracellular pigments will be correlated with changes in rates of mixed-function oxidation. (2) Study the effect of altered nutritional status and selected inducing agents on rates of conjugation of the oxidized products of the model substrates mentioned above in isolated prfused tissues. Rates of conjugation via various pathways will be compared with specific enzymes and measured concentrations of activated intermediates such as UDP glucuronic acid and PAPS to define rate-limiting factors for conjugation in intact tissues. We will attempt to enhance rates of conjugation of electrophiles generated from selected carcinogens in intact tissues. (3) Explore factors regulating mixed-function oxidation and conjugation in perfused livers of various strains of mice known to differ in their susceptibility to chemical carcinogens. Initial studies will involve DBA/2J and C57BL/6J which are well characterized in terms of their response to specific carcinogens and inducing agents. (4) Examine the influence of various nutritional factors and inducing agents on the microheterogeneity of enzymes associated with the formation of reduced cofactor, selected intermediates of energy metabolism and one or more components of the mixed-function oxidase system in liver using quantitative histochemical techniques.